The importance of postmenopausal estrogen replacement therapy in affording protection against the delayed neuronal death associated with cardiac arrest or cardiac surgery in women is a topic of intense research. Studies by this laboratory have shown that long-term treatment with estrogen at physiological levels ameliorates ischemia-induced hippocampal injury and prevents activation of apoptotic signaling cascades in the hippocampal CA1 (Jover et al., J. Neurosci 22:2115-2124, 2002). The objectives of the proposed research are to identify the cellular site(s) of action and the molecular mechanisms by which estrogen acts to block apoptotic cell death. Estrogens exert their genomic actions by interaction with the estrogen receptor ERa and ERbeta. The central hypothesis driving the proposed research is that estrogen acts via ERa and coordinate activation of IGF-I to block the onset of global ischemia-induced apoptotic cell death. Specific Aims are: 1) Identify the cellular targets that mediate the neuroprotective actions of estrogen on hippocampal neurons. Pharmacological studies will examine the ability of ER subtype-specific agonists to elicit protection and of broad-spectrum ER antagonists to abrogate protection by estrogen. Molecular genetic studies will examine the ability of estrogen to afford protection against hippocampal injury in mice with targeted deletions of the ERa or ERbeta gene. The 2-vessel occlusion model of global ischemia in mice is established in our laboratories (Oguro et al., J. Neurosci. 21:7534-7542, 2001). Additional experiments will assess whether ischemia alters the expression of ERa and/or ER-beta in the hippocampus. 2) Determine whether estrogen acts via coordinate activation of ERa and IGF-IR to protect hippocampal neurons, and if so, whether protection is mediated by the PI3K (phosphotidylinositol 3-kinase)/Akt (protein kinase B) and/or the MAPK (mitogen-activated protein kinase) signaling cascades. Specific experiments will assess 1) the ability of IGF-I receptor antagonists to abrogate estrogen-induced protection; 2) ability of IGF-I to mimic protection by estrogen; 3) ability of IGF-I to rescue neurons in Era-knockout, but not ER-beta-knockout, mice; 4) ability of Akt inhibitors to abrogate estrogen/IGF-I protection; and 5) ability of MAPK inhibitors to block estrogen/IGF-I protection. 3) Examine the apoptotic death cascades that are triggered by global ischemia including the Fas/Forkhead and caspase death cascades, and the ability of estrogen to intervene in these cascades. It is well established that the risk of cardiac arrest in women increases after menopause, a state characterized by hypoestrogenemia. Likewise, premenopausal women show an overall reduced incidence of cerebral stroke compared with men of equivalent ages. These studies are expected to shed light on the role of estrogen in affording protection against global ischemia arising after cardiac arrest or cardiac surgery in women. [unreadable] [unreadable]